D2-40 lymphatic marker for detecting lymphatic invasion in thin to intermediate thickness melanomas: Association with sentinel lymph node status and prognostic value-A retrospective case study
ABSTRACT Sentinel lymph node (SLN) status is the greatest prognostic factor of morbidity in melanoma. D2-40 antibody specifically marks lymphatic endothelium and has been used for identifying lymphatic invasion (LI) in multiple cancers.
We sought to determine the relationship between melanoma lymphatic invasion (as detected using D2-40 on primary melanoma biopsies/excisions) and the presence or absence of melanoma in subsequent SLN biopsy.
We retrospectively evaluated LI using D2-40 on primary biopsies/excisions from patients with thin to intermediate thickness (Breslow thickness: ≤2.0 mm) melanomas, who underwent lymphatic mapping and SLN biopsy, and whose SLN status was known. Sixty-four cases met the criteria and were available for analysis. We analyzed patient age, patient sex, mitotic rate, ulceration, tumor depth, and D2-40 detected LI as predictors of SLN status.
Lymphatic invasion detection increased from 3.1% using hematoxylin and eosin only to 21.9% using D2-40. Twelve of 14 patients with D2-40 LI were SLN positive (positive predictive value, 85.7%). D2-40 LI was detected in the primary biopsy specimen of 12 of 18 patients with a positive SLN (sensitivity 66.7%). Of 50 patients without D2-40 LI, 44 were SLN negative (negative predictive value, 88.0%). Of 46 SLN-negative patients, 44 did not have D2-40 LI (specificity, 95.7%).
Results are retrospective and limited to SLN biopsy performed at one institution.
On univariate and multivariate analysis, D2-40-detected LI was the most significant predictor of SLN status. D2-40 antibody staining to detect lymphatic invasion should be incorporated in routine melanoma biopsy evaluation.
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ABSTRACT: The aims of this study were to investigate the role of vascular invasion (blood and lymphatic), vessel density and the presence of tumour-associated macrophages as prognostic markers in 202 cutaneous melanoma patients. Sections of primary melanoma were stained with lymphatic-specific antibody D2-40 to assess lymphatic vessel invasion and density in intratumoural and peritumoural areas; an antibody against endothelial marker CD34 was used to determine blood vessel invasion and density, and an antibody against CD68 was used to determine macrophage counts. Immunohistochemically determined vascular invasion (combined blood and lymphatic) was compared with that determined using haematoxylin and eosin (H&E) staining. The use of immunohistochemistry increased detection of vascular invasion from 8-30% of patients, and histological exam of H&E-stained tissue was associated with a false positive rate of 64%. Lymphatic vessel invasion occurred at a much higher frequency than blood vessel invasion (27 and 4% of patients, respectively). Although immunohistochemically detected vessel invasion was significantly associated with histological markers of adverse prognosis, such as increased Breslow thickness, ulceration and mitotic rate (all P<0.001), no associations with relapse-free or overall survival were observed. High macrophage counts were significantly associated with markers of aggressive disease, such as Breslow thickness, ulceration and mitotic rate (P<0.001, P<0.001, P=0.005, respectively), and lymphatic vessel invasion and high microvessel density (P=0.002 and P=0.003, respectively). These results suggest that vascular invasion is more accurately detected using immunohistochemistry and occurs predominantly via lymphatic vessels. The association of vessel characteristics with histological characteristics of the primary melanoma provides evidence for their biological importance in melanoma, but that they were not associated with clinical outcome attests to the value of existing histological prognostic biomarkers. We note that a high macrophage count may be associated with neovascularisation and primary tumour growth, and may also promote invasion through lymphatic vessels.Modern Pathology 11/2011; 25(4):493-504. DOI:10.1038/modpathol.2011.182 · 6.36 Impact Factor
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ABSTRACT: Extracellular matrix protein 1 (ECM1) and vascular endothelial growth factor-C (VEGF-C) are secretory glycoproteins that are associated with lymphangiogenesis; these proteins could, therefore, play important roles in the lymphatic dissemination of tumors. However, very little is known about their potential roles in lymphangiogenesis. The aim of this study was to investigate whether correlations exist between ECM1 and VEGF-C in human breast cancer, lymphangiogenesis, and the clinicopathological characteristics of the disease. ECM1 and VEGF-C mRNA and protein expression levels in 41 patients were investigated using real-time reverse transcriptase polymerase chain reaction (RT-PCR), or immunohistochemical (IHC) staining of breast cancer tissue, matched noncancerous breast epithelial tissues, and suspicious metastatic axillary lymph nodes. D2-40 labelled lymph vessels and lymphatic microvessel density (LMVD) were counted. Correlations between ECM1 or VEGF-C protein expression levels, LMVD, and clinicopathological parameters were statistically tested. The rate of ECM1 positive staining in breast cancer tissues was higher (31/41, 75.6%) than that in the corresponding epithelial tissues (4/41, 9.8%, P < 0.001) and lymph nodes (13/41, 31.7%, P < 0.001). Similarly, the VEGF-C expression rate in cancer specimens was higher (33/41, 80.5%) than in epithelial tissues (19/41, 46.3%, P < 0.01) or lymph nodes (15/41, 36.6%, P < 0.01). Higher ECM1 and VEGF-C mRNA expression levels were also detected in the tumor tissues, compared to the non-cancerous tissue types or lymph nodes (P < 0.05). ECM1 protein expression was positively correlated with the estrogen receptor status (P < 0.05) and LMVD (P < 0.05). LMVD in the ECM1- and VEGF-C-positive tumor specimens was higher than that in the tissue types with negative staining (P < 0.05). Both ECM1 and VEGF-C were overexpressed in breast cancer tissue samples. ECM1 expression was positively correlated with estrogen responsiveness and the metastatic properties of breast cancer. We conclude, therefore, that ECM1 and VEGF-C may have a synergistic effect on lymphangiogenesis to facilitate lymphatic metastasis of breast cancer.BMC Cancer 01/2012; 12:47. DOI:10.1186/1471-2407-12-47 · 3.32 Impact Factor
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ABSTRACT: BACKGROUND: Approximately 4% of cutaneous squamous cell carcinomas (cSCCs) develop lymphatic metastases. The value of lymphatic endothelial markers to enhance the detection of lymphatic tumor invasion in cSCC has not been assessed previously. OBJECTIVE: We sought to evaluate the use of the antibody D2-40, a podoplanin immunohistochemical marker, to identify tumor lymph vessel invasion in cSCC and to assess its expression in tumor cells. METHODS: This was a retrospective case-control study. A series of 101 cSCC, including 51 cases that developed lymphatic metastatic spread (metastasizing cSCC [MSCC]) and 50 cases that resolved definitely after surgical excision (non-MSCC) were included in the study. Lymph vessel invasion using D2-40 was evaluated on all primary biopsy specimens. The percentage of tumor cells showing D2-40 positivity and intensity scoring were recorded. All the immunohistochemical findings were correlated with the clinicopathological features. RESULTS: Lymph vessel invasion was observed in 8% of non-MSCCs and in 25.5% of MSCCs (P = .031). D2-40 expression was significantly increased, both in intensity (odds ratio 4.42 for intensity ++/+++) and in area (odds ratio 2.29 for area >10%), in MSCC when compared with non-MSCC. Interestingly, almost half (49%) of the MSCC had moderate to intense D2-40 positivity compared with 16% of non-MSCC. D2-40 immunohistochemical expression was increased in tumors with an infiltrative pattern of extension. In the multivariate analysis, histologically poorly differentiated tumors, recurrent lesions, and cSCC showing D2-40 overexpression (in intensity) were significantly associated with lymphatic metastases development (odds ratios 15.67, 14.72, and 6.07, respectively). LIMITATIONS: This was a retrospective study. CONCLUSION: The expression of podoplanin associates with high metastatic risk in cSCC.Journal of the American Academy of Dermatology 04/2012; 67(6). DOI:10.1016/j.jaad.2012.03.007 · 5.00 Impact Factor